HONORS BIOLOGY LABORATORY ACTIVITIES – PHOTOSYNTHESIS
This laboratory activity is a demonstration of your understanding of the process of photosynthesis. You
have to write a complete lab report after you completed the procedure. The lab has 3 parts. For your
lab report, you will have a common title, abstract, background for all procedures.
PART I: DESIGNING AN EXPERIMENT BY USING THE FLOATING DISK METHOD
The biology behind the prodedure:
Leaf disks float, normally. When the air spaces are infiltrated with solution the overall density of the leaf
disk increases and the disk sinks. The infiltration solution includes a small amount of Sodium
bicarbonate. Bicarbonate ion serves as the carbon source for photosynthesis. As photosynthesis
proceeds oxygen is released into the interior of the leaf which changes the buoyancy--causing the disks
to rise. Since cellular respiration is taking place at the same time, consuming oxygen, the rate that the
disks rise is an indirect measurement of the net rate of photosynthesis.
Materials:







Sodium bicarbonate (Baking soda)
Liquid SoapPlastic syringe (10 cc or larger)—remove any needle!
Leaf material
Hole punch
Plastic cups
Timer
Light source
Optional:



Buffer Solutions
Colored Cellophane or filters
Different types of plants
Procedure:


Prepare 300 ml of bicarbonate solution for each trial. The bicarbonate serves as an alternate
dissolved source of carbon dioxide for photosynthesis. Prepare a 0.2% solution. (This is not very
much it is only about 1/8 of a teaspoon of baking soda in 300 ml of water.)
Add 1 drop of dilute liquid soap to this solution. The soap wets the hydrophobic surface of the
leaf allowing the solution to be drawn into the leaf. It’s difficult to quantify this since liquid
soaps vary in concentration. Avoid suds. If your solution generates suds then dilute it with more
bicarbonate solution.







Cut 10 or more uniform leaf disks for each trial.
Single hole punches work well for this but you can also just cut similar sized little squares with
scissors.
Choice of the leaf material is perhaps the most critical aspect of this procedure. The leaf surface
should be smooth and not too thick. We will use fresh spinach. Avoid major veins.
Infiltrate the leaf disks with sodium bicarbonate solution.
Remove the piston or plunger and place the leaf disks into the syringe barrel. Replace the
plunger being careful not to crush the leaf disks. Push on the plunger until only a small volume
of air and leaf disk remain in the barrel (< 10%).
Pull a small volume of sodium bicarbonate solution into the syringe. Tap the syringe to suspend
the leaf disks in the solution.
Holding a finger over the syringe-opening, draw back on the plunger to create a vacuum. Hold
this vacuum for about 10 seconds. While holding the vacuum, swirl the leaf disks to suspend
them in the solution. Let off the vacuum. The bicarbonate solution will infiltrate the air spaces
in the leaf causing the disks to sink. You will probably have to repeat this procedure 2-3 times in
order to get the disks to sink. If you have difficulty getting your disks to sink after about 3
evacuations, it is usually because there is not enough soap in the solution. Add a few more
drops of soap.

Pour the disks and solution into a clear plastic cup. Add bicarbonate solution to a depth of
about 3 centimeters. Use the same depth for each trial. Shallower depths work just as well.

For a control infiltrate leaf disks with a solution of only water with a drop of soap--no
bicarbonate.
Place under the light source and start the timer. At the end of each minute, record the number
of floating disks. Then swirl the disks to dislodge any that are stuck against the sides of the cups.
Continue until all of the disks are floating.

The Design Part:
Your group needs to design an experiment to use either multiple different types of plants or use
different colored filters or intensities of light to determine the effect of one of these factors on the rate
of photosynthesis.
In your lab report:
For this part, you need to include all usual parts of the lab report (hypothesis, materials, procedure with
all variables– shorten what is in this packet but you are not allowed to cut it out and glue it into your lab
report, data table, graph, conclusion)
PART II: OBSERVING STOMATA ON SPINACH LEAVES
Introduction
Plants and animals both have a layer of tissue called the epidermal layer. Plants have special pores
called stomata to allow passage of material. The stomata pores are surrounded on both sides by
jellybean shaped cells called guard cells. Unlike other plant epidermal cells, the guard cells contain
chlorophyll to do photosynthesis. This allows the cells to expand/ contract to open or close the
stomata. Guard cells also close when dehydrated. This keeps water in the plant from escaping. The
opening or closing of guard cells can be viewed in a microscope by adding different water concentration
to the leaf tissue.
Most stomata are on the lower epidermis of the leaves on plants (bottom of the leaf). The number of
stomata on the epidermal surface can tell you a lot about a plant. Usually, a high concentration of
stomata indicates fast growth and wet climate. Lower concentrations of stomata indicate lower rates of
photosynthesis and growth or adaptations for dry weather.
Purpose:
To view and compare the stomata from the leaves of several species of plant
Materials:
3 leaves (1 from 3 different species), compound light microscope, 3 microscope slides, clear nail polish,
transparent tape
Procedure:
1. Obtain three leaves from different types of plants.
2. Paint a thick patch (at least one square centimeter) of clear nail polish on the underside of the
leaf surface being studied.
3. Allow the nail polish to dry completely.
4. Tape a piece of clear cellophane tape to the dried nail polish patch.
5. Gently peel the nail polish patch from the leaf by pulling on a corner of the tape and "peeling"
the fingernail polish off the leaf. This is the leaf impression you will examine.
6. Tape your peeled impression to a very clean microscope slide. Use scissors to trim away any
excess tape. Label the slide with plant name with a sharpie.
7. Examine the leaf impression under a light microscope at 400X.
8. Search for areas where there are numerous stomata, and where there are no dirt, thumb prints,
damaged areas, or large leaf veins. Draw the leaf surface with stomata.
9. Count all the stomata in one microscopic field. Record the number on your data table.
10. Repeat counts for at least two other distinct microscopic fields. Record all the counts. Determine
an average number per microscopic field.
11. From the average number/400X microscopic field, calculate the stomata per mm2 by multiplying
by 8.
12. Follow procedures 2 - 11 with the other leaves.
In your lab report:
(Complete the hypothesis, materials, procedure, data table, drawing, graph and conclusion sections)
PROCEDURE III: LEAF CHROMATOGRAPHY
Background Information:
Paper chromatography is a process that uses special filter paper to separate and identify the different
substances in a mixture. Chromatography means “to write with color.” The substances in the mixture
dissolve in the alcohol and move up the paper. The heavier substances move up the paper more slowly.
The lighter substances move up the paper more quickly. Heavy and light substances get separated from
one another on the paper.
Plants contain chlorophyll, a green pigment, as well as carotenoids, pigments that range in color from
red to orange to yellow.
Materials:
 Isopropyl alcohol
 ruler
 spinach leaf
 scissors
 chromatography paper
 pencil
 paperclip
 tape
 beaker
 colored pencils
Procedure:
1. Obtain a strip of chromatography paper.
2. Use a ruler to measure and draw a light pencil line 2-cm above the bottom of the paper strip.
3. Here is the tricky part! Place the edge of the spinach leaf over the pencil line and using the edge of a
coin gently press on the spinach leaf to create a single green line over the pencil line. You want this line
to be thin and concentrated with the pigment from the spinach leaf. Therefore, repeat this edging
process carefully about 3-4 times. Be sure not to press too hard or you will poke a hole through the
paper.
4. Carefully add isopropyl alcohol to the beaker until it reaches a depth of 1-cm in the beaker.
5. Tape the top of the paper strip to a pencil so that the end of the strip with the green line hangs down.
The pencil should be able to sit across the top of the beaker with the bottom of the paper strip just
touching the bottom of the beaker. Cut off any excess paper from the TOP of the strip if it is too long.
6. Lay the pencil across the top of the beaker with the paper strip extending into the alcohol. MAKE
SURE THAT THE LEVEL OF THE ALCOHOL IS BELOW THE GREEN LINE ON YOUR PAPER
STRIP! IF THE ALCOHOL IS GOING TO COVER THE GREEN LINE, POUR OUT SOME
ALCOHOL BEFORE YOU GET THE GREEN LINE WET!
8. Observe as the alcohol gets absorbed and travels up the paper by capillary action. This may take up to
20 minutes. Do not touch or move your experiment during this time.
9. When the alcohol has absorbed to approximately 1-cm below the pencil, you may remove the
pencil/paper strip from the beaker to dry on your counter. With a pencil, mark the distance the alcohol
has traveled on the paper, as well as the distance each pigment has traveled.
10. Using colored pencils, draw your results in the data table.
11. Using a ruler and the following formula, measure the Rf values of each pigment.
Since the fastest molecules will travel the greatest distance, or to the highest point along the strip, the
relative distances can be measured, and the flow rate (migration) of the molecules (Rf) can be
calculated by using the following formula:
Rf = Distance pigment traveled/ Distance solvent traveled
12. Include your distance traveled and Rf values in your data table.
13. Identify the pigments and last include that in your data table:
Pigment Identification:
Chlorophyll a= blue-green
Chlorophyll b = Olive green
Xanthophyll = yellow
Carotene = orange yellow
In your lab report:
Include background, materials, procedure, data table and conclusion.